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Overview Sensitivity Tuner --- Characterisation --- Modelling Colour Generators --- Carotenoids (Orange/Red) --- Melanin (Brown) --- Violacein (Purple/Green) The Future Safety

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Week 8

Monday

Wet Work

Threshold device

Transformed 72, 92, 94, and 95 in pSB3K3 into arabinose strain.

Melanin Biobrick

Did the last stage of PCR for the Melanin Biobrick creation. Fragments AB and CF were added with primers A and F and the Finnzymes enzyme, then run on a CYBR-safe gel. The results showed smears rather than bands. As the CF fragment was a little dodgy on the first extraction it was decided to re-extract and try again.

The CF band was reproduced and extracted in the afternoon, creating more CF fragment stock.

Dry Work

Tuesday

Wet Work

Threshold device

Successfully moved 70, 71, 72, 74, 91 into pSB3K3. That makes all 15 activator constructs yay!

Melanin Biobrick

Re-tried the PRC using the new CF fragment under the same PCR conditions. Still getting smeared bands.

Dry Work

Wednesday

Wet Work

Melanin Biobrick

Looked up the possible reasons for smeared bands after PCR, three main ones discovered:

• Too much template DNA
• Incorrect salt concentration
• Comtaminated PCR mix

Nanodropped the fragments and diluted AB 1/10 to get the recommended DNA concentration, and used new pipettes and Primer aliquots. Two tubes were made up using a new Finnzymes mix, and two using the old one.

The gel still showed smears! For all the samples.

Thursday

Friday